Influenza A virus (IAV) infections affect millions of people worldwide every year and cause serious mortality. Current treatment options are limited to viral strain-specific vaccination and are problematic due to antiviral drug resistance. There is an urgent need to identify novel host innate immune mechanisms providing broad range protection against influenza. Bronchial epithelial cells orchestrate an oxidative extracellular antimicrobial system present in the airway surface liquid consisting of the protein lactoperoxidase (LPO), the thiocyanate anion (SCN-) and hydrogen peroxide (H2O2). LPO oxidizes SCN- using H2O2 into hypothiocyanite (OSCN-) that has known in vitro antiviral effects. Dual oxidase 1 (Duox1), an NADPH oxidase highly expressed in bronchial epithelial cells, is the H2O2 source for the system. Our long-term goal is to determine whether the Duox1/H2O2/LPO/SCN- antiviral system could be manipulated in influenza infection for therapeutic purposes. The objective of this proposal is to determine the antiviral role of Duox1 against influenza. Our preliminary data show that 1) primary bronchial epithelial cells inactivate influenza in an H2O2/LPO/SCN--dependent manner, 2) Duox1-deficient mice are impaired in early lung clearance of IAV, and 3) LPO and SCN- added exogenously to human bronchial epithelial cells following IAV challenge improves viral clearance. Based on these data, our central hypothesis is that the Duox1/LPO/SCN- system inactivates extracellular influenza virions in a strain- independent manner and attenuates IAV infection and lung damage in the mouse lung. The rationale for the proposed research is that we need to characterize how powerful is this system in fighting influenza and how easily can it be manipulated. Our hypothesis will be tested by these specific aims: 1) Determine the mechanism of IAV inactivation by the Duox1/LPO/SCN- system in BECs, in vitro; 2) Determine the role of Duox1 in a murine model of IAV lung infection, in vivo; and 3) Determine whether in vivo therapeutic manipulation of the Duox1/H2O2/LPO/SCN- system attenuates IAV lung infection. It is anticipated that our aims will yield the following outcomes: 1) detailed description of the mechanism of Duox1 in the anti-influenza response of BECs; 2) establishing the in vivo relevance of Duox1 in fighting IAV; and 3) testing the first therapeutic approach to boost Duox1/H2O2/LPO/SCN- system for improved IAV clearance. Our innovative work shows that the H2O2/LPO/SCN- system inactivates influenza, uses a Duox1-deficient mouse strain for in vivo studies and suggests that promoting this system could attenuate influenza infection and lung damage. The significance of the outlined work relies in establishing the relevance of the Duox1/H2O2/LPO/SCN- system in fighting influenza. In summary, our proposed work will have a positive impact on the fields of airway epithelial biology and antiviral innate immune responses by identifying Duox1, as a novel, crucial weapon of the bronchial epithelium against influenza.